Toll-like receptors (TLRs) play a pivotal role in shaping then host immune response to a pathogen or a vaccine. Our understanding of the mechanisms by which this occurs, have arisen from explorations that probe the response of immune cells to a single TLR ligand. However, microbes and vaccines do not simply stimulate a single TLR, but rather stimulate combinations of different TLRs. Recent work by Lanzavecchia and others suggests that the combinatorial activation of multiple TLRs result in a synergistic activation of cytokine production by dendritic cells (DCs). The impact of this synergy on the adaptive immune response is poorly understood. In particular, there is little or no understanding of the innate immune mechanisms that affect critical variables of the B cell response, such as memory B cell generation, affinity maturation, and induction of neutralizing antibodies. Our preliminary data in mice, suggest that TLR ligands administered with an antigen, can elicit antigen-specific antibody responses. In particular, specific combinations of TLR ligands result in a synergistic induction of the antigen-specific antibody responses, and in the induction of high avidity antibodies. The precise mechanism by which this occurs is a mystery, and will be the major focus of this proposal, and will be achieved in the following aims: Aim 1: To determine whether combined stimulation with TLR4 + TLR7/8 ligands results in a synergistic activation of the germinal center pathway of memory B-cell differentiation Aim 2: To determine the cellular and molecular mechanism(s) by which TLR ligands act synergistically to stimulate antigen-specific B cell responses Aim 3: To determine the "quality" of antibody that is optimally suited for protection against specific category A-C agents The successful completion of these aims, will greatly facilitate the rational design of vaccines that stimulate optimally effective types of B cell responses against various pathogens.